The long-term goal is to determine 1) if mature astrocytes in intact brain exhibit membrane receptors for neuroactive substances and 2) if so, what events are regulated by such receptors. There is extensive evidence that astroglia isolated from immature brain and grown in culture express a variety of neurotransmitter receptor systems. However, there is little information concerning the functionality of these receptors on cultured cells and essentially no information regarding whether there is heterogeneity among astroglia with respect to receptor expression or whether astroglial receptor systems are expressed in mature brain. Studies from this laboratory indicate that there are at least two pharmacologically distinct populations of astroglia which can be distinguished on the basis of their expression of beta-adrenergic receptors (B-AR). These two populations of astroglia appear to represent separate lineages of astrocytes which have been termed type 1 astroglia (polygonal GF AP+ cells which express B-AR) and type 2 astroglia (process-bearing GFAP+ cells which lack B-AR). Our more recent studies suggest that type 1 astroglia may be subdivided on the basis of their expression of alpha-1 adrenergic (A1-AR) receptors. These results were obtained using a novel technique which enables us to examine receptors via receptor autoradiography on immunocytochemically identified cells in primary cell culture. Using this technique it is possible to examine the presence of receptors by individual cells, in a manner analogous to the study of neurons using electrophysiology, within a large population of cells. As a result, factors such as sample purity and yield are no longer limiting in analyzing receptor expression among nonexcitable brain cells. This research proposal will expand our analysis of receptor expression on single, immunocytochemically identified brain cells by using established fluorescent indicators of cellular calcium, pH, and membrane potential to detect receptor mediated responses. In addition, we propose to study receptor mediated changes in cyclic AMP levels in single cells using immunocytochemical techniques. A number of questions concerning the pharmacological properties of astrocytes will be addressed using these techniques. First, is there heterogeneity among astroglia with respect to receptor expression? Second, do interactions occur among neural cells which influence the expression of astroglial receptor? Third, do astrocytes either isolated from mature brain or present in the intact brain exhibit a similar set of receptors as cultured astroglia. Results from thse studies should provide important information concerning the capacity of astrocytes to respond to neuroactive substances released in brain and, as a result, provide valuable information concerning the function of astrocytes in brain.